■ Abstract Apoptosis is a genetically programmed, morphologically distinct form of cell death that can be triggered by a variety of physiological and pathological stimuli. Studies performed over the past 10 years have demonstrated that proteases play critical roles in initiation and execution of this process. The caspases, a family of cysteine-dependent aspartate-directed proteases, are prominent among the death proteases. Caspases are synthesized as relatively inactive zymogens that become activated by scaffold-mediated transactivation or by cleavage via upstream proteases in an intracellular cascade. Regulation of caspase activation and activity occurs at several different levels: ( a) Zymogen gene transcription is regulated; ( b) antiapoptotic members of the Bcl-2 family and other cellular polypeptides block proximity-induced activation of certain procaspases; and ( c) certain cellular inhibitor of apoptosis proteins (cIAPs) can bind to and inhibit active caspases. Once activated, caspases cleave a variety of intracellular polypeptides, including major structural elements of the cytoplasm and nucleus, components of the DNA repair machinery, and a number of protein kinases. Collectively, these scissions disrupt survival pathways and disassemble important architectural components of the cell, contributing to the stereotypic morphological and biochemical changes that characterize apoptotic cell death.

Mammalian caspases: structure ,a ctivation ,s ubstrates, and functions during apoptosis

Apoptosis plays an important role in development and maintenance of tissue homeostasis. Intensive efforts have been made to explore the molecular mechanisms of the apoptotic signaling pathways including the initiation, mediation, execution and regulation of apoptosis. Caspases are central effectors of apoptosis. Cells undergo apoptosis through two major pathways, namely the extrinsic pathway (death receptor pathway) or the intrinsic pathway (the mitochondrial pathway). Finally, the contents of dead cells are packaged into apoptotic bodies, which are recognized by neighboring cells or macrophages and cleared by phagocytosis. Cellular apoptosis is tightly controlled by a complex regulatory networks including balancing pro-survival signals. De-regulation of apoptosis may lead to pathological disorders such as developmental defects, autoimmune diseases, neurodegeneration or cancer. Increasing attention is being focused on alternative signaling pathways leading to cell death including necrosis, autophagy, and mitotic catastrophe. Understanding of cell death signaling pathways is relevant to understanding cancer and to developing more effective therapeutics.

Overview of cell death signaling pathways

The loss of vital cells within healthy tissues contributes to the development, progression and treatment outcomes of many human disorders, including neurological and infectious diseases as well as environmental and medical toxicities. Conversely, the abnormal survival and accumulation of damaged or superfluous cells drive prominent human pathologies such as cancers and autoimmune diseases. Apoptosis is an evolutionarily conserved cell death pathway that is responsible for the programmed culling of cells during normal eukaryotic development and maintenance of organismal homeostasis. This pathway is controlled by the BCL-2 family of proteins, which contains both pro-apoptotic and pro-survival members that balance the decision between cellular life and death. Recent insights into the dynamic interactions between BCL-2 family proteins and how they control apoptotic cell death in healthy and diseased cells have uncovered novel opportunities for therapeutic intervention. Importantly, the development of both positive and negative small-molecule modulators of apoptosis is now enabling researchers to translate the discoveries that have been made in the laboratory into clinical practice to positively impact human health.

Regulation of apoptosis in health and disease: the balancing act of BCL-2 family proteins

Apoptosis (programmed cell death) is a cellular self-destruction mechanism that is essential for a variety of biological events, such as developmental sculpturing, tissue homeostasis, and the removal of unwanted cells. Mitochondria play a crucial role in regulating cell death. Ca2+ has long been recognized as a participant in apoptotic pathways. Mitochondria are known to modulate and synchronize Ca2+ signaling. Massive accumulation of Ca2+ in the mitochondria leads to apoptosis. The Ca2+ dynamics of ER and mitochondria appear to be modulated by the Bcl-2 family proteins, key factors involved in apoptosis. The number and morphology of mitochondria are precisely controlled through mitochondrial fusion and fission process by numerous mitochondria-shaping proteins. Mitochondrial fission accompanies apoptotic cell death and appears to be important for progression of the apoptotic pathway. Here, we highlight and discuss the role of mitochondrial calcium handling and mitochondrial fusion and fission machinery in apoptosis.

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The role of mitochondria in apoptosis

For over three decades, a mainstay and goal of clinical oncology has been the development of therapies promoting the effective elimination of cancer cells by apoptosis. This programmed cell death process is mediated by several signalling pathways (referred to as intrinsic and extrinsic) triggered by multiple factors, including cellular stress, DNA damage and immune surveillance. The interaction of apoptosis pathways with other signalling mechanisms can also affect cell death. The clinical translation of effective pro-apoptotic agents involves drug discovery studies (addressing the bioavailability, stability, tumour penetration, toxicity profile in non-malignant tissues, drug interactions and off-target effects) as well as an understanding of tumour biology (including heterogeneity and evolution of resistant clones). While tumour cell death can result in response to therapy, the selection, growth and dissemination of resistant cells can ultimately be fatal. In this Review, we present the main apoptosis pathways and other signalling pathways that interact with them, and discuss actionable molecular targets, therapeutic agents in clinical translation and known mechanisms of resistance to these agents.

Targeting apoptosis in cancer therapy

https://arrow.tudublin.ie/cgi/viewcontent.cgi?article=1039&context=scschbioart

Regulation and measurement of oxidative stress in apoptosis.

Control of mitochondrial integrity by Bcl-2 family members and caspase-independent cell death.

1. Whey and Whey Products Whey can be defined in a very general sense as "the liquid remaining after removal of casein from milk". Casein can be separated from milk in a variety of ways resulting in the production of a number of different types of whey, each of which can be identified on the basis of the method of casein removal. From an industrial viewpoint there are two principal types of whey (IDF, 1978; Nielsen, 1974); sweet and acid wheys. These are by-products of cheese or rennet casein and acid casein manufacture, respectively. Acidification may be by direct addition of mineral acid or by in situ production of acid by added starter bacteria Sweet whey has a minimum pH of 5.6 and acid whey a maximum pH of 5.1 (IDF, 1978). Whey composition varies depending on milk source, processing methods used and the cheese or casein type but typically it contains ~ 50% of the total solids of the original milk and includes lactose, proteins, minerals and vitamins. Data on whey composition include those by Roeper (1971), Cerbulis, Woychik and Wondolowski (1972), Josephson, Rizvi and Harper (1975), Glass and Hedrick (1976a, b) and Marshall (1982). Table 1 lists the average composition of some types of whey. Whey was traditionally regarded as a waste product and was generally disposed of as effluent or as an animal feed. However, it has now come to be regarded as a product whose valuable constituents can be constructively utilized by the food industry, where there has been increased recognition that the proteins, lactose and possibly even the salts in whey are valuable nutrients. Impetus for whey utilization is due to the increased use of functional proteins in processed foods and to the development of new technologies for the economic recovery of such protein from whey. Heating has direct relevance in the development of whey processing techniques and influences the functionality of whey protein products; it is a major technological treatment, either as a process per se or as part of a process. Heating induces changes in whey constituents, especially the proteins, which have a major influence on the efficiency of the process and on the quality of the product. Thus, heating has direct effects on both the structure and functionality of these proteins in food systems. An understanding of the effects of heat on proteins, i.e. denaturation and aggergation, might permit manipulation and control of these phenomena in order to permit greater product and process control and/or to develop new processes or products with predictable or tailor-made functional properties. For example, Richert, Morr and Cooney (1974) and Richert (1979) showed that controlled heating, to induce limited 43

Whey Proteins and their Thermal Denaturation - A Review

Light-induced photoreceptor apoptosis in vivo requires neuronal nitric-oxide synthase and guanylate cyclase activity and is caspase-3-independent.

Apoptosis is the mode of cell death in retinitis pigmentosa, a group of retinal degenerative disorders primarily affecting rod photoreceptors. Although caspases have been demonstrated to play a central role in many incidences of apoptosis, accumulating evidence suggests that they may not be required for all forms of apoptotic cell death. The present study examined the mechanism of cell death in two in vivo models of photoreceptor apoptosis: the retinal degeneration (rd) mouse, a naturally occurring mutant model, and N-methyl-N-nitrosourea-induced retinal degeneration. Specifically, we examined the activation status of caspase-9, -8, -7, -3, and -2 and determined the caspase requirements for cytochrome c release, DNA fragmentation, and apoptosis-associated proteolysis of specific caspase substrates. We show that apoptosis in both in vivo models is independent of caspase-9, -8, -7, -3, and -2 activation. DNA fragmentation occurs in the absence of caspase-mediated ICAD (inhibitor of caspase-activated DNase) proteolysis, suggesting that an alternative endonuclease is responsible for DNA cleavage in these models. Importantly, we show that apoptosome activation is prevented because of an absence of mitochondrial cytochrome c release. Experiments performed using a cell-free system indicate that cytochrome c-dependent proteolysis and activation of caspase-9 can be restored in a neonatal cell-free system. However, we found that cytochrome c-dependent proteolysis and activation of caspase-9 could not be restored in an adult cell-free system because of an age-related decrease in the expression of Apaf-1 in the normal developing mouse retina. In the rd mouse, however, this age-related downregulation of apoptotic proteins was not observed, highlighting a critical feature of this model and the prevention of cytochrome c release as an apical event in caspase-independent apoptosis in this system.

https://www.jneurosci.org/content/jneuro/23/13/5723.full.pdf

Maryanne Donovan

Papers

Regulation and measurement of oxidative stress in apoptosis.

Control of mitochondrial integrity by Bcl-2 family members and caspase-independent cell death.

Whey Proteins and their Thermal Denaturation - A Review

Light-induced photoreceptor apoptosis in vivo requires neuronal nitric-oxide synthase and guanylate cyclase activity and is caspase-3-independent.

Caspase-Independent Photoreceptor Apoptosis in Mouse Models of Retinal Degeneration